PC Spacer (photocleavable) is a non-nucleosidic molety that can be used as an intermediary to attach any available phosphoramidite modification at either end of an oligonucleotide through a UV photo-cleavable C3 spacer, as well as insert such a spacer internally. An example is the use of PC Spacer to incorporate a photo-cleavable 6-FAM tag onto the 5’-end of oligonucleotides immobilized on glass slides. These fluorescently-labeled oligo arrays were then UV irradiated in order to test the efficacy of photo-cleavage in removing the 6-FAM tag from these oligos, as part of developing sequencing-by-synthesis applications (1).
Cleavage Protocol
Cleavage occurs by irradiation with near-UV light (300-350 nm, complete cleavage occurs within 5 minutes. Try using a Black Ray XX-15 UV lamp (Ultraviolet Products Inc., San Gabriel, CA) at a distance of 15 cm (emission peak 365 nm, 300 nm cut-off, 1.1 mW intensity at~31 cm).
References
1. Olejnik, J., Krzymanska-Olejnik, E., Rothschild, K.J. Photocleavable aminotag phosphoramidites for 5’-termini DNA/RNA labeling. Nucleic Acids Res. (1998), 26: 3572-3576.
2. Olejnik, J., Ludemann, H-C., Olejnik, E.K, Berkenkamp, S., Hillenkamp, F., Rothschild, K.J. Photocleavable peptide-DNA conjugates: synthesis and applications to DNA analysis using MALDI-MS. Nucleic Acids Res. (1999), 27: 4626-4631.
3. Tang, X., Su, M., Yu, LiLi, Lv, C., Wang, J., Li, Z. Photomodulating RNA cleavage using photolabile circular antisense oligodeoxynucleotides.Nucleic Acids Res. (2002), 38: 3848-3855.
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